Comparison of Monoclonal Antibodies and Polymerase Chain Reaction Assays for the Typing of Isolates Belonging to the D and M Serotypes of Plum Pox Potyvirus

Abstract

Plum pox potyvirus (PPV) isolates may be divided into four groups separated by serological, molecular, and epidemiological differences. Monoclonal antibodies specific for the two major groups of isolates, represented by the D and M serotypes of the virus, have been obtained. Polymerase chain reaction (PCR)-based assays allowing the direct detection and differentiation of PPV isolates have also been developed. We now report on a large-scale comparison of these two typing approaches. The results obtained show an overall excellent correlation between the results obtained in indirect double-antibody sandwich enzyme-linked immunosorbent assay using PPV-D- and PPV-M-specific monoclonal antibodies and those derived from either specific PCR assays or restriction fragment length polymorphism analysis of PCR fragments. Without exception, all isolates reacting positively with the PPV-M-specific monoclonal antibody were found to belong to the M serotype using the PCR-based assays, while 51 out of 53 isolates recognized by the D-specific monoclonal antibodies belonged to the D serotype according to the PCR typing results. However, failure to react with a specific monoclonal antibody did not prove as effective a predictor of the serotype of the isolate analyzed. In a few cases, the results obtained with the various techniques diverged, indicating low level variability of the epitopes recognized by the serotype-specific monoclonal antibodies. Isolates belonging to the two minor groups of PPV (El Amar and Cherry) also gave divergent results, indicating that the current typing assays are not suited for the analysis of such isolates.

Strain identification and sequence variability of plum pox virus in Turkey

“…However, we are cautious about the rates of mixed infection in Ankara. We assume that most of the mixed infections identified by Elibüyük (2011) could be PPV-T, similar to the misinterpretation of PPV-T as PPV-M, because T isolates could also react to PPV-D-specific antibodies Candresse et al, 1998;Myrta et al, 1998). Other than D isolates in Ankara, PPV-D occurrence has not been reported in other regions in Turkey.…”

“…Sequence analysis is accepted as the most reliable method to identify the virus. Similar to other plant viruses, PPV has been mostly analyzed by sequencing the viral CP gene (Bousalem et al, 1994;Candresse et al 1998). Typing methods for the P3-6K1 region have also been developed, and the P3-6K1 region has been used for PPV variability studies, allowing PPV subgroup discrimination independent of the CP (Glasa et al, 2002).…”

Plum pox virus strains: Diversity and geographical distribution in Serbia

“…The most frequently targeted region in PPV analysis is the highly variable 5'-terminal part of the CP gene. Many strain-specific primers targeting this and other parts of the PPV genome have been developed for single or mixed infection detection (Candresse et al, 1995;Candresse et al, 1998;Nemchinov & Hadidi, 1998;Glasa et al, 2002b;Šubr et al, 2004;Glasa et al, 2005;Glasa et al, 2013). PPV strains can also be discriminated by PCR-RFLP analysis, but some isolates may be mistakenly classified.…”

Plant Pathogen Forensics: Capabilities, Needs, and Recommendations

“…Reasonably "universal" primers have been developed for some virus families, genera, and "species" targeted to taxon-specific sequences (35,91,150). RT-PCR clearly differentiates between some strains of common viruses: for example, between PPV strains D and M (27) and SMV strains G2 and G7 (131) The methods mentioned above are all limited by their reliance on a minute fraction of a taxon's many defining features. For nucleic acid-based methods, the accuracy of the comparison to some degree will be proportional to the length of the fragment (and/or the site) used in the analysis.…”